1. Field of the Invention
The present invention relates to an absorbance detection system in a lab-on-a-chip, and more particularly, to a high-efficiency, high-sensitivity absorbance detection system in which detection cells with long optical path lengths for higher detection sensitivity, micro-lenses for collimating light into the detection cells, and slits for preventing scattered light from entering detectors are fabricated and integrated in a lab-on-a-chip.
The present invention of an absorbance detection system in a lab-on-a-chip is capable of solving the problems of conventional absorbance detection systems in lab-on-a-chips and produces a 10 times greater detection sensitivity.
The invented detection system can utilize liquid- or solid-state waveguides or micro-light sources (lamp or laser) as well as optical fibers for light radiation.
The present invention has additional collimators including micro-lenses and slits arranged close to detection cells so that effective absorbance detection can be achieved using the detection cells with the 50 μm or greater optical pathlengths.
The present invention of an absorbance detection system in a lab-on-a-chip has collimators including micro-lenses for collimating divergent light from optical fibers and transmitting the collimated light to detection cells. Slits are built-in to prevent light that does not pass through the detection cell or scattered light from entering detectors so that detection sensitivity can be improved markedly with increased optical pathlength of the detection cell.
2. Description of the Related Art
A variety of analytical instruments such as capillary electrophoresis (CE), liquid chromatography, and gas chromatography are used to separate and analyze mixed compounds. In particular, CE and liquid chromatography have wide applications in conjunction with a variety of available detection methods, such as absorbance detection, fluorescence detection, electrochemical detection and others. The fluorescence detection method advantageously has a high detection sensitivity but needs fluorescence labels to be coupled to samples because self-luminescent substances rarely exist. The electrochemical method also has a high detection sensitivity but its application is limited to specific compounds. Whereas, the absorbance detection method can be applied to a wide range of analytes and does not need labeling of analytes for detection, and thus it has been the most popular detection method.
In the absorbance detection method based on Beer's law, the absorbance of a sample is proportional to the distance light passes through the sample, i.e., the optical pathlength, which is expressed by:A=ε×b×C  (1)where A is absorbance, ε is the molar extinction coefficient (L/mol·cm), b is the optical pathlength (cm), C is the molar concentration of the sample (mol/L).
Sensitivity of the absorption detection is usually poor in CE. This is because the capillary used for CE has a small inner diameter of 50–100 μm, and thus the optical pathlength is very short. In addition, because the capillary has a circular cross-section, a portion of light passes through the capillary, and thus the actual pathlength is smaller than the inner diameter of the capillary.
In an attempt to increase the sensitivity of absorbance detection in CE, detection cells with an extended optical pathlength have been developed. Typically, the use of a capillary having a rectangular cross-section or a U-shaped or Z-shaped detection cell has been suggested to increase the optical pathlength by 10–50 times the inner diameter of a common capillary.
In a lab-on-a-chip based CE system, the depth of micro-channels formed in a glass plate or plastic plate is as small as 10–30 μm, and thus the absorbance detection sensitivity in the microchip system is worse than that in CE. For this reason, an attempt to increase detection sensitivity by applying a U-shaped detection cell in a lab-on-a-chip system has been made. In this approach, optical fibers are arranged in front of and behind the detection cell, and light is radiated into the detection cell through an optical fiber and collected by another optical fiber for detection.
Lab-on-a-chip systems for chemical/biological analysis will be described briefly. Lab-on-a-chip systems fabricated by a micro-machining technique such as a photolithography technique used in the manufacture of semiconductor devices are referred to as chemical microprocessors including a variety of components (for sample pretreatment, injection, reaction, separation and detection) integrated in a glass, silicon, or plastic substrate of an area of several square centimeters. These lab-on-a-chip systems advantageously enable high-speed, high-efficiency, high-cost automated chemical/biological analysis to be carried out just on the one device.
In most lab-on-a-chip based analytical systems, migration and separation of a sample are performed by electroosmotic flow induced by the application of voltages to both ends of a microchannel filled with a sample solution. The microfluidics in a microchip can be controlled by applying high voltages and thus eliminating the use of a mechanical pump or valve. This has enabled the microchip to be fabricated to much smaller sizes than other commercially available analytical systems and at relatively low costs. In addition, a series of sample injection, migration, reaction, separation and detection processes can be performed continuously in a single lab-on-a-chip.
Although the lab-on-a-chip-based analytical systems described above are advantageous in that the consumption of sample and reagents is reduced and the analysis can be performed within a short period of time, they cannot be applied to the analysis of a variety of samples due to limited detection methods. So, fluorescence detection and electrochemical detection methods are commonly used for detection in a lab-on-a-chip. To compensate for the drawback of the lab-on-a-chip-based analytical systems and to extend its applications, a glass based lab-on-a-chip integrated with an absorption detection system using optical fibers was developed. In this system, a single mode optical fiber having a small numerical aperture and a small core diameter was used in order that almost all of the radiated light passes through a U-shaped detection cell. Light passed through a single mode optical fiber diverges conically at a predetermined angle. The diameter (w) of light from the single mode optical fiber is calculated by:w=d×(0.65+1.619/V15+2.879/V6)  (2)where d is the diameter of the optical fiber, and V=d×π×NA/λ where NA is the numerical aperture of the optical fiber, and λ is the wavelength of radiated light.
With this type of a conventional absorption detection system in a lab-on-a-chip, an optical fiber having a core diameter of 3 μm, a cladding diameter of 125 μm, and an NA of 0.1 is used, and 488-nm light is radiated from an argon ion laser. The diameter of light from this optical fiber, which can be calculated by formula (2) above, is 3.93 μm. Here, the divergence angle (θ) of light is calculated by:θ=arc sin(NA/n)  (3)where n is the refractive index of a medium through which light transmits (n=1.33 for water, n=1.52 for glass). The diameter (w′) of divergent light at a distance (X) from the medium is calculated by:w′=w+2X tan θ  (4)
As an example, assuming that light from a single mode optical fiber passes through a detection cell filled with water and having a length of 150 μm, the divergent light from the detection cell, which can be calculated by the formulae above, has a diameter of about 27 μm. If the detection cell has a length of 500 μm, the divergent light from the detection cell has a diameter of about 80 μm. Therefore, when designing a U-shaped detection cell having a depth of 25 μm and a width of 50 μm for a lab-on-a-chip using a single mode optical fiber, the length of the U-shaped detection cell is determined to be no larger than 150 μm to allow almost all the light radiated through the single mode optical fiber to pass through the U-shaped detection cell. As a result, the conventional absorbance detection system in a lab-on-a-chip showed an increase in detection sensitivity by only 3–4 times of that of a detection method in an non-extended detection cell.
In addition, because the conventional absorbance detection system in a lab-on-a-chip is fabricated in glass, it is difficult to fabricate, is time consuming and has low reproducibility. Also, the detection cell (channel) of the absorbance detection system has a circular cross-section and thus generates a serious light scattering problem.